Biology subjects

Butt, U.

Publications and source records attributed to Butt, U..

4 recordsLinked to original sources

bioRxiv2022

SHARPIN serine 146 phosphorylation mediates ARP2/3 interaction, cancer cell invasion, and metastasis

The adaptor protein SHARPIN is involved in a number of cellular processes and promotes cancer progression and metastasis. However, how the choice between different functions of SHARPIN is post-translationally regulated is unclear. Here we have characterized SHARPIN phosphorylation by mass spectrometry and in vitro kinase assay. Focusing on two uncharacterized phosphorylation sites, serine 131 and 146, in the unstructured linker region of SHARPIN, we demonstrate their role in SHARPIN-ARP2/3 complex interaction, whereas they play no role in integrin inhibition or LUBAC activation. Consistent with its novel role in ARP2/3 regulation, serine 146 (S146) phosphorylation of SHARPIN promoted lamellopodia formation. Notably, CRISPR-Cas9 mediated knockout of SHARPIN abrogated three-dimensional (3D) invasion of several cancer cell lines. The 3D invasion of cancer cells was rescued by overexpression of the wild-type SHARPIN, but not by SHARPIN S146A mutant, identifying S146 as an invasion promoting phosphorylation switch. Finally, we demonstrate that inhibition of phosphorylation at S146 significantly reduces the in vivo metastasis in the zebrafish model. Collectively, these results demonstrate that SHARPIN S146 phosphorylation constitutes a single functional determinant of cancer cell invasion both in vitro and in vivo.

cancer biology↗

bioRxiv2021

Ovarian cancers with low CIP2A tumor expression constitute an APR-246 sensitive disease subtype

Identification of ovarian cancer (OvCa) patient subpopulations with increased sensitivity to targeted therapies could offer significant clinical benefit. We report that 22% of the high grade OvCa tumors at diagnosis express CIP2A oncoprotein at low levels. CIP2Alow OvCa tumors have significantly lower likelihood of disease relapse after standard chemotherapy, but yet a portion of relapsed tumors retain their CIP2Alow phenotype. We further discover that reactive oxygen species (ROS) inducing compound APR-246 (PRIMA-1Met/Eprenetapopt), currently in clinical development, preferentially kill CIP2Alow OvCa cells across multiple chemotherapy resistant cell lines. Consistent with CIP2Alow OvCa subtype in humans, CIP2A is dispensable for development of MISIIR-TAg-driven mouse OvCa tumors. Nevertheless, CIP2A deficient OvCa tumor cells from MISIIR-TAg mice displayed APR-246 hypersensitivity both in vitro and in vivo. Mechanistically, the lack of CIP2A expression hypersensitizes the OvCa cells to APR-246 by inhibition of NF-kB activity. Accordingly, combination of APR-246 and Nf-kB inhibitor compounds strongly synergized in killing of CIP2A positive OvCa cells. Collectively, we discover low CIP2A expression as a vulnerability for APR-246 in OvCa. The results warrant consideration of clinical testing of APR-246 for CIP2Alow OvCa tumor subtype patients, and reveal CIP2A as a candidate APR-246 combination therapy target.

cancer biology↗

bioRxiv2020

CIP2A interacts with TopBP1 and is selectively essential for DNA damage-induced basal-like breast cancer tumorigenesis

Despite saturated genetic profiling of breast cancers, oncogenic drivers for the clinically challenging basal-like breast cancer (BLBC) subtype are still poorly understood. Here, we demonstrate that CIP2A is selectively essential for DNA damage-induced initiation of mouse BLBC tumors, but not of other cancer types. Mechanistically, CIP2A was discovered genome-widely the closest functional homologue for DNA-damage proteins TopBP1, RHNO, POLQ, NBN and PARP1. CIP2A directly interacts with the ATR-activation domain of TopBP1, and dampens both, chromatin binding of TopBP1 and RAD51, and G2/M checkpoint in DNA-damaged cells. CIP2A also drives BLBC-associated proliferative MYC and E2F1 signaling. Consistently with high DNA-damage response activity BLBCs, and CIP2As novel role in checkpoint signaling, CIP2A was found essential for DNA-damaged, and BRCA-mutant BLBC cells. Selective role for CIP2A as BLBC driver was supported by association of high CIP2A expression with poor patient prognosis only in BLBC, but not in other breast cancer types. Therapeutically, small molecule reactivators of PP2A (SMAPs) phenocopy CIP2A-dependent DNA damage response, and inhibit in vivo growth of patient-derived BLBC xenograft. In summary, we discover sub-type selective essential role for CIP2A in BLBC initiation and maintenance that can be explained by its newly discovered association with DNA-damage response, coordinated with regulation of proliferative signaling. The results also identify therapeutic strategy for CIP2A-dependent BLBCs.

cancer biology↗

bioRxiv2019

Protein Interactome of the Cancerous Inhibitor of Protein Phosphatase 2A (CIP2A) in Th17 Cells

Cancerous inhibitor of protein phosphatase 2A (CIP2A) is involved in immune response, cancer progression, and in Alzheimers disease. However, an understanding of the mechanistic basis of its function in this wide spectrum of physiological and pathological processes is limited due to its poorly characterized interaction networks. Here we present the first systematic characterization of the CIP2A interactome by affinity-purification mass spectrometry combined with validation by selected reaction monitoring targeted mass spectrometry (SRM-MS) analysis in Th17 cells. In addition to the known regulatory subunit of protein phosphatase PP2A, the catalytic subunit of protein PP2A was found to be interacting with CIP2A. Furthermore, the regulatory (PPP1R18, and PPP1R12A) and catalytic (PPP1CA) subunits of phosphatase PP1 were identified among the top novel CIP2A interactors. Evaluation of the ontologies associated with the proteins in this interactome revealed that they were linked with RNA metabolic processing and splicing, protein traffic, cytoskeleton regulation and ubiquitin-mediated protein degradation processes. Taken together, this network of protein-protein interactions will be important for understanding and further exploring the biological processes and mechanisms regulated by CIP2A both in physiological and pathological conditions.\n\nHighlights{blacksquare} The first characterisation of the CIP2A interactome in Th17 cells.\n{blacksquare}Key interactions were validated by targeted SRM-MS proteomics, western blot and confocal microscopy.\n{blacksquare}Pathway analysis of the interactome revealed interrelationships with proteins across a broad range of processes, in particular associated with mRNA processing.

systems biology↗